The development of hemoglobin-based oxygen carriers has focused on oxygen delivery for use in medical therapies such as transfusions and the production of blood products. Hemoglobin-based oxygen carriers can be used to prevent or treat hypoxia resulting from blood loss (e.g, from acute hemorrhage or during surgical operations), from anemia (e.g., pernicious anemia or sickle cell anemia), or from shock (e.g, volume deficiency shock, anaphylactic shock, septic shock or allergic shock).
Existing hemoglobin-based oxygen carriers include perfluorochemicals, synthesized hemoglobin analogues, liposome-encapsulated hemoglobin, chemically-modified hemoglobin, and hemoglobin-based oxygen carriers in which the hemoglobin molecules are cross-linked. Preparation of hemoglobin-based oxygen carriers includes several purification steps. In order to remove plasma proteins from whole blood, a process of microfiltration is used to wash the cells of whole blood. The cell washing operation removes plasma proteins from bovine whole blood using diafiltration over a 0.2 μm microfiltration membrane with isotonic saline/citrate solution. Diafiltration is a continuous filtration operation in which saline/citrate solution is added to the filter retentate to maintain a volume in the recirculation tank. The blood solution is recirculated across the filter and the filtrate, containing the plasma proteins, is sent to waste.
Washing the blood solution using filtration results in highly variable processing times which adversely effect product throughput. Additionally, extended cell washing process times could lead to growth of unacceptable levels of bioburden and to cell lysis, thereby further reducing process yield.